The study of molecular neurobiology has developed with a view to implementing specific, effective therapeutic strategies to treat neural debilitation. Classically, three technical approaches have been followed to the present. The first, comprising the cloning and sequencing of novel cDNAs, seeks to develop specific genes that may be subsequently introduced in vivo to mediate and thereby achieve physiological alterations. The second strategy of molecular hybridization for the purpose of studying expression and regulation of specific messenger RNAs, likewise seeks to promote modifications through transcription. By contrast, the third strategy, involving direct manipulation of neuronal expression in vivo, seeks to actively alter neuronal cell physiology.
Two strategies that have been investigated in the direct approach involve the development of a transgenic vehicle, and efforts to deliver specific genes by incorporation within a viral vector that is delivered directly to the neuronal population of interest for in vivo expression. The former approach has been limited somewhat by corresponding limitation in the choice of species that are amenable to transgenic manipulations. Also, the transgenic approach has failed to provide the necessary specificity and selectively that is requisite for effective use. Tissue-specific promoters have been applied to this effect and have achieved some level of success [Forss-Petter et al. (1990) NEURON., 5:187-197; Oberdick et al. (1990) SCIENCE, 248:223-226]. Even with this degree of selectively, however, expression in all cell types in which a given promoter is active is still unavoidable.
The use of viral constructs is the subject of extensive study, and particularly, with respect to the exploration of such a construct involving a defective herpes simplex viral vector based on a plasmid called an amplicon [Kaplitt et al. (1991) MOL. CELL. NEUROSCI., 2:320-330; Spaete et al. (1982) CELL, 30:295-304; Geller et al. (1988) SCIENCE, 241:1667-1669]. Herpes simplex virus type 1 (HSV1) has been favored because of its broad host and tissue ranges. Several investigators including the present inventor have experimented with the incorporation into HSV1 vectors, and particularly defective viral vectors, various foreign genes for direct neuronal introduction in vivo. While such efforts have been pursued, and in particular, efforts with the lac Z gene of E. coli [Geller et al. supra], none of the vectors developed and used have ever resulted in stable expression, by which is meant expression in excess of two weeks of a gene introduced in a defective HSV vector.
Other investigators have pursued gene transfer in vivo by use of an HSV whole viral vector [see generally, Breakefield et al. (1991) NEW BIOL., 3(3):203-218]. Successful genetic transfer in vivo has been achieved with these vectors [Dobson et al. (1990) NEURON, 5:353-360; Chiocca et al. (1990) NEW BIOL., 2:739-746; Fink et al. (1992) HUMAN GENE THER., 3:11-19]. While all of the noted investigators observed expression after introduction with the whole viral vector, expression ceased after two weeks in two instances and extended to five weeks and then ceased in the third [Dobson et al., supra]. In this latter connection, Dobson et al. noted that the lac Z gene was placed under the control of the viral promoter, and that the normal viral gene does not express during HSV latency, so that the cessation of expression after five weeks was attributed to cessation of promoter activity upon the entry of the viral DNA into a latent state.
As the potential value of stable and long-term expression of foreign genes in target neuronal populations and, particularly, in the brain, holds great promise for both diagnostic and therapeutic applications, it is clear that a need exists for the development of such a system and corresponding vector as will achieve such stable and long-term expression. It is to the achievement of this objective that the present invention is therefore directed.
The citation or identification of any reference in this application shall not be construed as an admission that such reference is available as prior art to the present invention.